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Background: Booster doses could play a major role in no responders or low responders to primary hepatitis B (HB) vaccine. Planed time point for hepatitis A vaccination in China provides a good opportunity to carry out HB booster dose by using combined hepatitis A and B vaccine.

Methods: A randomized, double-blinded clinical trial was conducted to compare the immunogenicity and safety of toddlers 18–24 months of age receiving 3 different vaccination regimens: 2 doses of inactivated hepatitis A vaccine (group 1), 1 dose of inactivated hepatitis A vaccine plus 1 dose of combined hepatitis A and B vaccine (group 2) or 2 doses of combined hepatitis A and B vaccine (group 3).

Results: All 3 groups showed 100% seroprotection for antihepatitis A virus antibody after vaccination. Seroprotection rate for anti-HB antibody before vaccination ranged from 79.5% to 92.9% in the 3 groups. After second inoculation, anti-HBs seroprotection increased from 92.9% to 100% in group 2 with postvaccination geometric mean concentration (GMC) of 2258.3 mIU/mL and from 79.5% to 98.9% in group 3 with postvaccination GMC of 2055.3 mIU/mL. The adverse events were not statistically different among groups (P = 0.345).

Conclusions: Combined hepatitis A and B vaccine could stimulate high level of both antihepatitis A virus and anti-HBs antibodies and not increase adverse events, providing a new choice for HB booster.

From the *Hunan Center for Disease Control and Prevention, Changsha, Hunan province, China; †Sinovac Biotech Ltd., Beijing, China; and ‡Chaling Center for Disease Control and Prevention, Zhuzhou, Hunan province, China.

Although hepatitis A and B vaccine are in place for many years and the incidence of acute hepatitis A and B has continued to fall since the 1990s, hepatitis virus still poses severe public health burden globally.1 It is estimated that there are more than 2 billion individuals exposed to hepatitis B (HB) virus and 250 million are chronically infected. HB is the cause of more than 600,000 annual deaths due to cirrhosis and hepatocellular carcinoma.2 World Health Organization estimated more than one million acute hepatitis A cases annually.3 In 2006, there were an estimated 15,000 cases of acute hepatitis A and 13,000 cases of HB in the United States.4 The incidence of viral hepatitis is around 100 per 100 thousand in China, with hepatitis A and B taking over 75%.5

HB vaccine has been shown to be highly efficient in preventing HB. Recombinant HB vaccine, which gradually replaces plasma-derived HB vaccine, has been launched since 1986, conducting 3 or 4 doses scheme starting with 1 monovalent vaccine on birth date.6 The primary 3-dose vaccine series showed protective antibody concentrations in most of healthy infants, children and young adults. Unfortunately, 5%–10% of healthy individuals and 40%–50% of hemodialysis patients do not develop an adequate immune response against the vaccine.7 A Chinese study found that 0.99% newborns did not respond to the vaccination(defined as antibody level <10 mIU/mL) and 11.39% had low level (between 10 and 100 mIU/mL) who still faced great risk of acquiring HB virus.8 Studies have shown that booster doses could play a major role in these no responders or low responders.7,9–11 However, it is not common to give booster dose based on serostatus by testing every infant after primary vaccination. Furthermore, the interval for booster vaccinations against HB virus (HBV) is still undecided.12 Appropriate timing for booster needs further studies.

Hepatitis A vaccine has been incorporated into routine national vaccination program in many countries worldwide. The inactivated hepatitis A vaccine conducts a 2-dose schedule starting at 18 months with 6 months apart in China.13 Combined hepatitis A and B vaccine, composing of inactivated hepatitis A vaccine and recombinant HB vaccine, launched in 2005 in China, which can prevent hepatitis A and B infection simultaneously and can reduce the number of needles and frequency of vaccination.14 With this product, the scheduled time for hepatitis A vaccination could serve as a good timing for HB booster vaccination.

Previous studies have proved the safety, immunogenicity and tolerability of the combined hepatitis A and B vaccine.4,15–17 Some experts argued that combined hepatitis A and B vaccine could elicit better immune response compared to HB monovalent vaccine, which may be due to the positive bystander effect of hepatitis A vaccine.18,19 Until now few study has been conducted to show the feasibility and effectiveness of HB booster vaccination strategy using combined hepatitis A and B vaccine. Here, we designed a double-blinded trial in toddler to compare the immunogenicity and safety between different immunization regimes with inactivated hepatitis A vaccine and combined hepatitis A and B vaccine, with a purpose of displaying the performance of the potential new HB booster strategy and providing evidence for policy makers.

MATERIALS AND METHODS

Study Design and Subjects

This randomized, double-blinded trial was conducted in Chaling County, Hunan province of China. This study was approved by the Ethics Committee of Hunan Center for Disease Control and Prevention before enrollment. The clinical trials were conducted in accordance with the principles of the Declaration of Helsinki, the standards of Good Clinical Practice (as defined by the International Conference on Harmonization) and the Chinese regulatory requirements. Before participation, written informed consent was obtained from each volunteer’s parents or guardian. The trial was registered with the ClinicalTrial.gov number NCT02445703.

A total of 301 toddlers 18–24 months of age who had not received inoculation of hepatitis A vaccine but finished primary dose schedule of HB vaccine were involved in this study. The toddlers were randomly assigned into 3 groups with about 100 each, receiving 2 doses of inactivated hepatitis A vaccine (group 1), 1 dose inactivated hepatitis A vaccine plus 1 dose combined hepatitis A and B vaccine (group 2) or 2 doses of combined hepatitis A and B vaccine (group 3). Zero- and 6-month schedule was chosen for all 3 groups. Blood samples were collected before inoculation and 1 month after the second dose.

Vaccines

Both the inactivated hepatitis A vaccine and the combined hepatitis A and B vaccine were produced by Sinovac and licensed by Chinese Food and Drug Administration. The inactivated hepatitis A vaccine was produced with strain TZ84 of hepatitis A virus (HAV), which was cultivated in 2BS human fetal lung diploid fibroblast, then harvested, purified by chromatography, inactivated by formalin and adsorbed onto aluminum hydroxide, containing 250 U of HAV as antigen (lot number: 201308046). The combined hepatitis A and B vaccine was made on the basis of hepatitis A vaccine, by mixing with alum-adjuvant recombinant HB virus surface antigen expressed in beer yeast, composing of 250 U HAV and 5 µg HB virus surface antigen (lot number: 201307017). Both vaccines were packaged in syringes (0.5 mL per vial).

Safety Assessment

Subjects were observed for 30 minutes after injection for any adverse events. The parents or guardians of the participants were asked to fill out diary cards that listed injection-site adverse reactions (eg, pain, redness and swelling) and systemic adverse reactions (eg, fever, irritability and loss of appetite) in the first 3 days. Adverse events were noted and recorded through weekly visit and report between 4 and 30 days. Serious adverse events were collected throughout the trial. Adverse reactions were graded according to the standard guideline published by Chinese Food and Drug Administration and National Institute of Allergy and Infectious Diseases of National Institutes of Health.

Statistical Analysis

Adverse event rates, seroprotection rates and geometric mean concentrations (GMCs) for both vaccines were analyzed. A χ2 test was used to compare adverse event rates and seroprotection rates based on antibody titers, and one-way analysis of variance was used to compare GMCs among the 3 vaccine groups. SPSS version 15.0 software was used to perform the analyses, and a P < 0.05(2 sided) was considered statistically significant.

RESULTS

Study Population

Between May 14 and 17, 2014, a total of 301 subjects were enrolled and received 1 dose of investigational vaccines. After 6 months, 272 children were available for inoculation of the second dose (90.3% of all subjects). Blood samples after full course of vaccination from 252 participants (83.7%) were collected for immunogenicity test. Information on study enrollment and randomization is shown in Figure 1. The 3 groups were well balanced in terms of mean age, height, weight and sex.

Immunogenicity

Seroprotection rates and GMCs of anti-HAV IgG before inoculation and 1 month after second inoculation are shown in Table 1. Before inoculation, no significant difference was shown for both seroprotection rate and GMC in all 3 groups (P = 0.865 and 0.742, respectively). All three groups showed 100% seroprotection rate 1 month after second inoculation, while significant difference was shown for both the GMC and the increase of GMC with P < 0.001. Subjects receiving 2 doses of inactivated hepatitis A vaccine showed highest GMC and increase of GMC, whereas subjects receiving 2 doses of combined hepatitis A and B vaccine showed lowest GMC and increase of GMC with P < 0.001 for all comparisons.

Seroprotection rates and GMCs of anti-HBs IgG before inoculation and 1 month after second inoculation are shown in Table 2. Seroprotection rate for anti-HBs antibody before vaccination ranged from 79.5% to 92.9% in the 3 groups. Before inoculation, no significant difference was shown for GMC in 3 groups, whereas there was a significant difference in terms of seroprotection rate among the 3 groups (P = 0.029) with lowest rate in subjects receiving 2 doses of combined hepatitis A and B vaccine. After second inoculation, anti-HBs seroprotection increased from 92.9% to 100% in group 2 with postvaccination GMC of 2258.3 mIU/mL and from 79.5% to 98.9% in group 3 with postvaccination GMC of 2055.3 mIU/mL. However, GMC in group 1 dropped almost a half from 79.0 to 40.5 mIU/mL and seroprotection rate decreased from 88.6% to 77.2%.

Safety

During the entire study period, adverse events were observed in 71 of 301 subjects (23.6%), including 46 grade 1 events (15.3%), 24 grade 2 events (8.0%) and 1 grade 3 event (0.3%). The adverse events rate was 28.9%, 20.4% and 22.5% in group 1, 2 and 3, respectively, with no statistically significant difference among groups (P = 0.345). Furthermore, no significant difference in the adverse events rates were demonstrated between the inactivated hepatitis A vaccine and the combined hepatitis A and B vaccine for both first and second inoculation with P = 0.729 and 0.630, respectively.

DISCUSSION

This study was performed to evaluate the efficacy and safety of the inactivated hepatitis A vaccine and the combined hepatitis A and B vaccine with different schedule and to provide evidence to decide best immunization regime for the 2 vaccines. Results indicated that 1 month after second inoculation, subjects in all 3 groups with negative anti-HAV antibody showed 100% seroprotection with a GMC more than 3000 mIU/mL, which was 150 times higher than the cutoff level. We also found that subjects receiving at least 1 dose of combined hepatitis A and B vaccine showed more than 98% seroprotection of anti-HBs antibody with GMC more than 2000 mIU/mL, which was 200 times higher than the cutoff level. This study demonstrated that the immunization effect of vaccination strategy with combined hepatitis A and HB vaccine was successful both in providing protection against HAV and in reinforcing protection against HB virus, similar to other studies using combined hepatitis A and B vaccine.20,21 However, to our knowledge, this study is rare in that it explored the opportunity and possibility to fully or partially replace the routine hepatitis A vaccine with combined A and B vaccine at the scheduled time point for hepatitis A vaccination.

Our study found that the bivalent presentation was less potent than the monovalent with respect to the titer of antibody induced against hepatitis A. This suggested that there was some kind of immunosuppression, although not a big issue since the seroprotection was still 100%. The immunosuppression could also be seen in other combined vaccines, such as DTaP-Hib-IPV and measles, mumps and rubella vaccine. The possible reasons include epitopic suppression, antigen excess and change of immunoreaction.22 However, the mechanism of inducing lower hepatitis A antibody by our hepatitis A and B is still not clear, and some studies are under way.

In this study, hepatitis A and B did not show dose–effect relationship for anti-HBs GMC after vaccination. Group receiving 2 doses of combined hepatitis A and B vaccine even had lower GMC of anti-HBs compared to the group receiving 1 dose. However, this might be attributed to the lower GMC of 2 doses group at baseline. If we look at the GMC increase, 2 doses group still performed better than 1 dose group.

Our study reinforced the fact that booster dose was effective for those with no response or low response to primary HB vaccine.10,23,24 In this study, there were 24 subjects with anti-HBs IgG level below 10 mIU/mL in group 2 and 3 before inoculation. Twenty-three of them developed immune response after either 1 or 2 doses combined hepatitis A and B.

Our study again raised the question of whether a booster of HB vaccine is needed.25 First, between 7.1% and 20.9% subjects in the 3 groups were seronegative for anti-HBs IgG before inoculation. Second, subjects in group 1 immunized with 2 doses of inactivated hepatitis A vaccine demonstrated decrease of anti-HBs antibody seroprotection rate from 86.6% to 77.2% and decline of GMC from 79.5 to 40.5 mIU/mL during the 7 months period of this study. Although many researchers insisted that it was not necessary for HB booster when a complete primary vaccination course was offered with a theory that anamnestic response could be initiated when human body faced invasion from HB virus,26,27 no responders or low responders were not considered in such theory.28

This study was subject to a few limitations. First, we did not investigate the persistence of anti-HAV and anti-HBs antibody; however, it may not be an issue according to quite a few relevant studies,29,30 which provided a solid basis for the understanding of the immunity to hepatitis vaccine antigens. Even so, the persistence of the investigational vaccine still needs future studies to confirm. Furthermore, we found that vaccination strategy with combined A and B vaccine was effective, safe and efficient; however, economic value of such strategy was not provided in this study. Previous cost-economic study has showed good value of combined A and B vaccine in healthcare workers.31 In the future, such health economic studies are needed for decision making and policy recommendations. Finally, there were 29 and 49 subjects who missed inoculation of the second dose and taking blood samples after full course vaccination, respectively, which might reduce the statistical power. However, comparisons between subjects in and out of the cohort showed there was no statistical difference with respect to age, gender, height and weight. So we think these missing subjects might not affect the overall conclusions.

In conclusion, providing children 16–28 months of age with inoculation of 1 or 2 doses of combined hepatitis A and B vaccine could stimulate both high level of anti-HAV and anti-HBs antibodies and not increase adverse events. This study suggested a potential HB booster vaccination strategy using combined hepatitis A and B vaccine and provided decision makers with a new choice of vaccine selection at the scheduled time point for hepatitis A vaccination.